Hi, I was wondering if a blot (western blot) can be kept in a fridge at 4C for a few days after secondary incubation? If so then should it be kept in TBST? I heard that over exposure with TBST will wash away the secondary, is that true?
Our scanner (Typhoon) broke down and it may take days before the technician could fix it. I tried using x-ray films but unfortunately (we haven't used x-ray films in ages) all of our developer and fixer have expired (wasted one blot trying).
Hello Elena
yes you can keep the blot but whether dry or in TBST you will indeed lose signal - from both primary and secondary - and in some cases non specific blocking sites will be unmasked to. Indeed, storing blots @ 4C in TBST is a slow way of stripping them before reprobing
Thus, wrap your blot in saran wrap and keep in a DRY environment. When your typhoon is back up and running then re probe with secondary and develop
If you incubate in TBST instead you will have to effectively start again, i.e. block for 1 hour; probe with primary antibody; secondary then develop, especially if your gene is not a high copy number and especially too if your blot is quantitative
Hope that helps
Good point by Nadia
it is the detergent that strips membranes
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